This project seeks an understanding at the molecular level of cystine storage disease or cystinosis, an inherited disease of children which invariably results in uremic death early in life. Since skin fibroblasts in tissue culture from these patients are known to have 100 times the normal cystine content, use of these cells can obviate the obstacle to biochemical investigation due to the unavailability of cystinotic tissue for study. Continuous culture of these cells provides a model system for application of isotopic and enzymologic techniques as well as ultramicroscopic analysis to the unanswered problem of why cystine accumulates in these cells and how this accumulation results in cellular damage. The influx and efflux rates of S35 labeled cystine, cysteine and methionine as well as the intracellular metabolic conversions will be studied in order to delineate the pathways of sulfur amino acid metabolism in normal and cystinotic cells. Combined with a kinetic isotope approach there will be an investigation of the activity of enzymes involved in sulfur amino acid metabolism. Radioautographic and electron microscopic analysis will attempt to delineate intracellular localization of stored cystine and the etiology of structural pathology. Studies of lysosomal enzyme function will focus on the nature of cellular cystine toxicity.